Estimation of Human Chronological Age from Buccal Swab Samples through a DNA Methylation Analysis Approach of a Five-Locus Multiple Regression Model.

Recent advancements in forensic genetics have facilitated the extraction of additional characteristics from unidentified samples. This study delves into the predictive potential of a five-gene (ELOVL2, FHL2, KLF14, C1orf132, and TRIM59) methylation rate analysis for human age estimation using buccal swabs collected from 60 Italian volunteers. The methylation levels of specific CpG sites in the five genes were analyzed through bisulfite conversion, single-base extension, and capillary electrophoresis. A multivariate linear regression model was crafted on the training set, then the test set was employed to validate the predictive model. The multivariate predictive model revealed a mean absolute deviation of 3.49 years in the test set of our sample. While limitations include a modest sample size, the study provides valuable insights into the potential of buccal swab-based age prediction, aiding in criminal investigations where accurate age determination is crucial. Our results also highlight that it is necessary to investigate the effectiveness of predictive models specific to biological tissues and individual populations, since models already proven effective for other populations or different tissues did not show the same effectiveness in our study.

An mRNA Profiling Study of Vaginal Swabs from Pre- and Postmenopausal Women.

Body fluid identification by means of mRNA profiling provides valuable supplementary information in forensic investigations. In particular, the detection of vaginal mucosa mRNA markers is highly relevant in sexual assault cases. Although the vagina undergoes characteristic age-related physiological changes over a lifetime, few studies have evaluated the efficacy of vaginal mRNA markers in women of different ages. In this multicentric study, a 19-plex mRNA profiling assay including vaginal-specific markers (CYP2B7P1, MUC4, MYOZ1) was tested in a collection of 6-20-month-old vaginal swabs obtained from pre- (n = 84) and postmenopausal (n = 55) female volunteer donors. Overall, participating laboratories were able to correctly identify ~85% of samples as vaginal mucosa by mRNA profiling. The assay's success rate did not differ between the two age groups and was not affected by the time interval between swab collection and RNA analysis. MYOZ1 resulted a less sensitive vaginal marker compared to MUC4 and CYP2B7P1. A significant relative increase in the contribution to the total amplification signal was observed for MUC4, compared to CYP2B7P1 and MYOZ1, in postmenopausal women. Observation of other body fluids and tissues different from vaginal mucosa was also evaluated in connection to information on previous sexual activity and menstrual cycle phase at the time of sampling.

Forensic Age Estimation through a DNA Methylation-Based Age Prediction Model in the Italian Population: A Pilot Study.

DNA methylation is one of the epigenetic marks which has been studied intensively in recent years for age predicting purposes in the forensic area. In order to integrate age prediction into routine forensic workflow, the purpose of this study was to standardize and optimize a DNA methylation-based protocol tailored to the Italian context. A previously published protocol and age-predictive method was implemented for the analysis of 84 blood samples originating from Central Italy. The study here presented is based on the Single Base Extension method, considering five genes: ELOVL2, FHL2, KLF14, C1orf132, now identified as MIR29B2C, and TRIM59. The precise and specific steps consist of DNA extraction and quantification, bisulfite conversion, amplification of converted DNA, first purification, single base extension, second purification, capillary electrophoresis, and analysis of the results to train and test the tool. The prediction error obtained, expressed as mean absolute deviation, showed a value of 3.12 years in the training set and 3.01 years in the test set. Given that population-based differences in DNA methylation patterns have been previously reported in the literature, it would be useful to further improve the study implementing additional samples representative of the entire Italian population.

Touch DNA Sampling Methods: Efficacy Evaluation and Systematic Review.

Collection and interpretation of "touch DNA" from crime scenes represent crucial steps during criminal investigations, with clear consequences in courtrooms. Although the main aspects of this type of evidence have been extensively studied, some controversial issues remain. For instance, there is no conclusive evidence indicating which sampling method results in the highest rate of biological material recovery. Thus, this study aimed to describe the actual considerations on touch DNA and to compare three different sampling procedures, which were "single-swab", "double-swab", and "other methods" (i.e., cutting out, adhesive tape, FTA® paper scraping), based on the experimental results published in the recent literature. The data analysis performed shows the higher efficiency of the single-swab method in DNA recovery in a wide variety of experimental settings. On the contrary, the double-swab technique and other methods do not seem to improve recovery rates. Despite the apparent discrepancy with previous research, these results underline certain limitations inherent to the sampling procedures investigated. The application of this information to forensic investigations and laboratories could improve operative standard procedures and enhance this almost fundamental investigative tool's probative value.

Comparison of the Allelic Alterations between InDel and STR Markers in Tumoral Tissues Used for Forensic Purposes.

Background and objectives: Over the last two decades, human DNA identification and kinship tests have been conducted mainly through the analysis of short tandem repeats (STRs). However, other types of markers, such as insertion/deletion polymorphisms (InDels), may be required when DNA is highly degraded. In forensic genetics, tumor samples may sometimes be used in some cases of human DNA identification and in paternity tests. Nevertheless, tumor genomic instability related to forensic DNA markers should be considered in forensic analyses since it can compromise genotype attribution. Therefore, it is useful to know what impact tumor transformation may have on the forensic interpretation of the results obtained from the analysis of these polymorphisms. Materials and Methods: The aim of this study was to investigate the genomic instability of InDels and STRs through the analysis of 55 markers in healthy tissue and tumor samples (hepatic, gastric, breast, and colorectal cancer) in 66 patients. The evaluation of genomic instability was performed comparing InDel and STR genotypes of tumor samples with those of their healthy counterparts. Results: With regard to STRs, colorectal cancer was found to be the tumor type affected by the highest number of mutations, whereas in the case of InDels the amount of genetic mutations turned out to be independent of the tumor type. However, the phenomena of genomic instability, such as loss of heterozygosity (LOH) and microsatellite instability (MSI), seem to affect InDels more than STRs hampering genotype attribution. Conclusion: We suggest that the use of STRs rather than InDels could be more suitable in forensic genotyping analyses given that InDels seem to be more affected than STRs by mutation events capable of compromising genotype attribution.

The Role of DNA Degradation in the Estimation of Post-Mortem Interval: A Systematic Review of the Current Literature.

The determination of the post-mortal interval (PMI) is an extremely discussed topic in the literature and of deep forensic interest, for which various types of methods have been proposed. The aim of the manuscript is to provide a review of the studies on the post-mortem DNA degradation used for estimating PMI. This review has been performed following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses and the PRISMA Guidelines. Several analytical techniques have been proposed to analyse the post-mortem DNA degradation in order to use it to estimate the PMI. Studies focused mainly on animal models and on particular tissues. The results have been mixed: while on the one hand literature data in this field have confirmed that in the post-mortem several degradation processes involve nucleic acids, on the other hand some fundamental aspects are still little explored: the influence of ante and post-mortem factors on DNA degradation, the feasibility and applicability of a multiparametric mathematical model that takes into account DNA degradation and the definition of one or more target organs in order to standardize the results on human cases under standard conditions.

Analysis of RNA in the estimation of post-mortem interval: a review of current evidence.

Post-mortem interval (PMI) determination is one of the most important issues in forensic sciences. In the past, forensic scientists provided different approaches (physical, chemical, and entomological) for the estimation of PMI without success.However, advances in molecular biology over the last two decades have allowed us to assess the time-dependent degradation of biological markers (e.g., proteins, DNA, and RNA). Thus, the aim of the manuscript is to provide a review of the recent progress in the estimation of PMI using molecular biology methods, mainly focusing on the potential usefulness of RNA markers. To this end, 29 studies have been systematically reviewed, each one chosen according to specific inclusion/exclusion criteria. The selected studies evaluated the contribution of endogenous reference genes in different biological samples in order to determine the PMI based on post-mortem RNA degradation as a function of other influencing factors such as time, cause of death, and environmental conditions.

Collecting sexual assault history and forensic evidence from adult women in the emergency department: a retrospective study.

BACKGROUND: The objective of this retrospective study was to examine the discrepancy between information derived from written medical reports and the results of forensic DNA analyses on swabs collected from the victims in 122 cases of alleged sexual assault treated at the Emergency Department of Padua Hospital. The examination of discrepant results has proved useful to support a broader application of sexual assault management, particularly during the taking of case history. METHODS: The Laboratory of Forensic Genetics of Padua University have processed samples from 122 sexual assault cases over a period of 5 years. RESULTS: Of the 103 cases in which the victim reported a penetration and ejaculation, only 67 (55% of all the samples) correlated with positive feedback match from the laboratory. In 36 cases in which the patient reported penetration with ejaculation, no male DNA was found in the samples collected. Therefore, there was a total of 41 cases in which the patient's report were not supported by laboratory data. In the remaining ten cases, which had an ambiguous history, 3 tested positively for the presence of male DNA. CONCLUSIONS: To avoid discrepancies between the medical reporting and reconstruction of sex crimes, it is crucial to deploy strategies which focus not only on the technical aspects of evidence collection, but also on how the victim's story is recorded; such efforts could lead to better management of sexual assault victims, and to a strengthened legal impact of forensic evidence and of crime reconstruction.

Certification ISO 9001 in clinical ethics consultation for improving quality and safety in healthcare.

This paper refers to the quality management process of the Laboratory of Clinical Bioethics (LCB) of the University of Padua (Italy), which has obtained the quality certification to ISO 9001:2008, as a Clinical Ethics Support Service. Its activities consist mainly in clinical ethics consultations and training services, addressed to those who are called to decisions with ethical implications in the clinical setting, proposing a structured approach to identify and analyze the ethical issues that may loom in the relationships between health professionals and patients, and participating in their solution. The expected benefits of the application of ISO 9001 were mainly the following: to formalize the procedure adopted for clinical ethics consultation and training, to obtain a controlled management of documents, information and data, to ensure and demonstrate the quality of the provided activities and to make methods and organization publicly available. The main results which have been achieved with the 'quality management project' are summarized as follows: the enunciation of LCB Mission and Quality Policy; the drafting of the procedure by which clinical ethics consultation is provided; the formalization of members' skills and the adoption of relevant process and outcome indicators. Our experience may be useful in promoting accountability for the quality of ethics consultation services. We consider the certification process as a tool for transparent and reliable management of one of the most critical tasks in the current context of healthcare, motivating similar facilities to undertake similar pathways, with the aim to provide quality control on their activities.

The Importance of Distinguishing Menstrual and Peripheral Blood in Forensic Casework: A Case Report.

This report presents a case of an 18-year-old woman found unconscious by police officers outside a club suspected to be victim of a sexual assault. She was not able to give information about the dynamics of the assault, since she was under the effect of a high amount of alcohol. She affirmed to be virgin.The forensic genetic analyses were performed on vaginal and rectal swabs as well as bloody-like traces present on the slip. DNA and RNA analysis were performed using a multiplex for 15 autosomal short tandem repeat markers and of 19 primers specific for different tissues, respectively.No male DNA was identified on vaginal and rectal swabs and on blood spots on the underwear. Blood, vaginal, and skin markers were scored observed in the samples obtained from slip traces and vaginal swab.The forensic genetic analysis supports the proposition that the victim had a sexual intercourse, in the absence of male biological material, in a case where the victim, altered by high blood alcohol levels, was unable to provide information.

Hospitalized hunger-striking prisoners: the role of ethics consultations.

We refer to hospitalized convicted hunger strikers in Padua Hospital who decided to fast for specific reasons, often demanding, to be heard by the judge, to complain about the existing custodial situation or to claim unjust treatment. The medical ethics of hunger strikers are debated because the use of force feeding by physicians is widely condemned as unethical, but courts, in Italy, sometimes order to transfer the convicted person to hospital and oblige healthcare practitioners to perform forcible feeding. This can engender a profound insecurity for the physicians taking action on the one hand, while preventing patients from fully availing themselves of this principle of self-determination on the other. Physicians are mainly concerned about how to manage this situation and they may request ethical consultation. When it comes to managing hospitalized hunger strikers, the ethics consultant may be able to facilitate the relationship between physicians and hunger strikers, enhance the latter's trust in the former, ensuring that strikers are aware of the risks associated with their fasting, and helping them to arrive of their own free will at the right decision concerning their behavior and their demands.

Discovering misattributed paternity in genetic counselling: different ethical perspectives in two countries.

Misattributed paternity or 'false' paternity is when a man is wrongly thought, by himself and possibly by others, to be the biological father of a child. Nowadays, because of the progression of genetics and genomics the possibility of finding misattributed paternity during familial genetic testing has increased. In contrast to other medical information, which pertains primarily to individuals, information obtained by genetic testing and/or pedigree analysis necessarily has implications for other biologically related members in the family. Disclosing or not a misattributed paternity has a number of different biological and social consequences for the people involved. Such an issue presents important ethical and deontological challenges. The debate centres on whether or not to inform the family and, particularly, whom in the family, about the possibility that misattributed paternity might be discovered incidentally, and whether or not it is the duty of the healthcare professional (HCP) to disclose the results and to whom. In this paper, we consider the different perspectives and reported problems, and analyse their cultural, ethical and legal dimensions. We compare the position of HCPs from an Italian and British point of view, particularly their role in genetic counselling. We discuss whether the Oviedo Convention of the Council of Europe (1997) can be seen as a basis for enriching the debate.

Effect of dactyloscopic powders on DNA profiling from enhanced fingerprints: results from an experimental study.

We conducted a study on the effect of fingerprint enhancement methods on subsequent short tandem repeat profiling. First, we performed a study typing blood traces deposited on 5 different surfaces, treated with 8 types of dactyloscopic powders. Three different DNA extraction methods were used. Subsequently, we analyzed latent fingerprints on the same 5 surfaces enhanced with the 8 different powders used in the first part of the study. This study has demonstrated that DNA profiling can be performed on fingerprints left on different substrates, and the substrate will affect the amount of DNA that can be recovered for DNA typing. In the first phase of the study, a profile was obtained in 92% of the 120 samples analyzed; in the second part, in 55% of the 80 samples analyzed, we obtained a profile complete in 32.5% of the cases. From the results obtained, it seems that the powders used in latent fingerprints enhancement, rather than having a direct inhibitory effect on extraction and amplification of DNA, may cause partial degradation of DNA, reducing the efficiency of amplification reaction. It should not be forgotten that these results were obtained under laboratory conditions, and in real caseworks, there may still be different problems involved.

Biobanking research on oncological residual material: a framework between the rights of the individual and the interest of society.

BACKGROUND: The tissue biobanking of specific biological residual materials, which constitutes a useful resource for medical/scientific research, has raised some ethical issues, such as the need to define which kind of consent is applicable for biological residual materials biobanks. DISCUSSION: Biobank research cannot be conducted without considering arguments for obtaining the donors' consent: in this paper we discuss to what extent consent in biobank research on oncological residual materials has to be required, and what type of consent would be appropriate in this context, considering the ethical principles of donation, solidarity, protection of the donors' rights and the requirements of scientific progress. Regarding the relationship between informed consent and tissue collection, storage and research, we have focused on two possible choices related to the treatment of data and samples in the biobank: irreversible and reversible anonymization of the samples, distinguishing between biobank research on residual materials for which obtaining consent is necessary and justified, and biobank research for which it is not. The procedures involve different approaches and possible solutions that we will seek to define. The consent for clinical research reported in the Helsinki Declaration regards research involving human beings and for this reason it is subordinate to specific and detailed information on the research projects. SUMMARY: An important ethical aspect in regard to the role of Biobanks is encouraging sample donation. For donors, seeing human samples being kept rather than discarded, and seeing them become useful for research highlights the importance of the human body and improves the attitude towards donation. This process might also facilitate the giving of informed consent more willingly, and with greater trust.

Ethical issues in DNA identification of human biological material from mass disasters.

Each mass disaster has its own characteristics and will involve a different approach, so the safeguarding and collection of forensic evidence have to be considered as part of the field response procedure. DNA typing has played a more prominent role in the identification of human remains, and particularly so for highly decomposed and fragmented remains. Although the ultimate goal is to obtain the identification, the specific context of each application of human identity testing has its specific problems, ranging from technical approach, through statistical interpretation, to ethical issues. The preparedness plan of the forensic genetics laboratory needs to include policies for family notification, long-term sample storage, and data archiving. For this reason, DNA sample collection and a strategy for DNA-based victim identification needs to be part of the preparedness plan. In this paper, the authors seek to define three of these ethical aspects: (1) the humanitarian importance of identification; (2) resource allocation in the victims' DNA identification; and (3) the secondary use for research of the samples initially collected for identification purposes. DNA analysis for the purpose of identifying victims of mass disasters has complex implications that demand much more rigorous examination than they have received until now.

Digital Biobanking and Big Data as a New Research Tool: A Position Paper.

Big data analytics in medicine is driving significant change, as it offers vital information for improving functions, developing cutting-edge solutions and overcoming inefficiencies. With the right archiving and analysis tools, all players in the healthcare system, from hospitals to patients and from medical personnel to the pharmaceutical industry, can yield numerous benefits. Therefore, to analyze and interpret these analytics effectively, so that they can be useful for the advancement of scientific knowledge, we require information sharing, specific skills, training, integration between all system players, unique infrastructures and security. All these characteristics will make it possible to establish and harmonize real big data biobanks, for which it will be appropriate to consider new forms of governance compared to those traditionally conceived for large-sample biobanks.

Lessons Learned from the COVID-19 Pandemic: A Survey-Based Study on a Sample of Italian Physicians' Opinions on Telemedicine.

Telemedicine was born out of the need to ensure clinical evaluation and personal care regardless of the physical presence of the healthcare professional nearby. Information technologies have been vital during the COVID-19 pandemic to ensure medical care and avoid the contagion between patients and clinicians. Accordingly, telecare services multiplied worldwide and gained paramount importance. The present work aims to collect field-based opinions about Telemedicine and ethics among Italian physicians. We developed a web-based questionnaire that was administered to Italian physicians from 1 May to 15 June 2022. The questionnaire was distributed as a link to Google Forms via social networks/instant messaging applications to groups of graduated and qualified physicians. A total of 180 physicians answered the questionnaire (with an age range from 25 to 68 years old). Physicians belonging to the medical area of expertise appear to more frequently use new technologies in comparison to other specialties. The vast majority believe that it is appropriate to use Telemedicine for monitoring and follow-up but not for evaluating a new patient. Concerns about changes in the physician-patient relationship, informed consent, digital barrier, and privacy and data protection also emerged. Finally, telehealth is thought to be a potential useful tool for the future by the majority of respondents but proper training for physicians is therefore needed.

Defining medical liability when artificial intelligence is applied on diagnostic algorithms: a systematic review.

Artificial intelligence (AI) in medicine is an increasingly studied and widespread phenomenon, applied in multiple clinical settings. Alongside its many potential advantages, such as easing clinicians' workload and improving diagnostic accuracy, the use of AI raises ethical and legal concerns, to which there is still no unanimous response. A systematic literature review on medical professional liability related to the use of AI-based diagnostic algorithms was conducted using the public electronic database PubMed selecting studies published from 2020 to 2023. The systematic review was performed according to 2020 PRISMA guidelines. The literature review highlights how the issue of liability in case of AI-related error and patient's damage has received growing attention in recent years. The application of AI and diagnostic algorithm moreover raises questions about the risks of using unrepresentative populations during the development and about the completeness of information given to the patient. Concerns about the impact on the fiduciary relationship between physician and patient and on the subject of empathy have also been raised. The use of AI in medical field and the application of diagnostic algorithms introduced a revolution in the doctor-patient relationship resulting in multiple possible medico-legal consequences. The regulatory framework on medical liability when AI is applied is therefore inadequate and requires urgent intervention, as there is no single and specific regulation governing the liability of various parties involved in the AI supply chain, nor on end-users. Greater attention should be paid to inherent risk in AI and the consequent need for regulations regarding product safety as well as the maintenance of minimum safety standards through appropriate updates.